Insulin resistance plays a major role in the pathogenesis of type 2 diabetes mellitus (T2DM) and it is the best predictor for the later development of the disease. The long-term objectives of this grant are to elucidate the cellular mechanisms of insulin resistance in skeletal muscle of healthy young lean insulin resistant offspring of parents with T2DM (IR offspring) and in healthy young insulin resistant obese individuals using a multidisciplinary team approach. This grant builds on our recent studies demonstrating a key role of intramyocellular lipid in the pathogenesis of skeletal muscle insulin resistance and a primary role of muscle insulin resistance in the pathogenesis of atherogenic dyslipidemia and non-alcoholic fatty liver disease in these individuals. Therefore, understanding the pathogenesis of insulin resistance in skeletal muscle of healthy young lean insulin resistant individuals and healthy insulin resistant obese individuals is the focus of this grant. Specifically we will examine: 1) whether modest weight loss can reverse insulin resistance in skeletal muscle by decreasing the elevated intramyocellular lipid (diacylglycerol) content and nPKC activity, independent of changes in muscle mitochondrial function and 2) whether alterations in basal and insulin stimulated rates of muscle mitochondrial pyruvate dehydrogenase (PDH)/tricarboxylic acid (TCA) flux contribute to intramyocellular lipid accumulation and insulin resistance in healthy lean IR offspring and healthy insulin resistant obese individuals as proposed by the Metabolic Inflexibility hypothesis of Kelley and coworkers. These questions will be addressed using novel state-of-the-art 13C/31P magnetic resonance spectroscopic and magnetic resonance imaging methods in combination with stable isotopes and GC and LC tandem mass spectrometry. Based on our strong preliminary data demonstrating no alterations in fasting or insulin stimulated PDH/TCA flux in IR offspring it is anticipated that the results of these studies will yield important new paradigm shifting insights into the pathogenesis of insulin resistance in skeletal muscle and that this will lead to the rational development of novel therapeutic targets to treat insulin resistance associated with obesity and T2DM. PUBLIC HEALTH RELEVANCE: Relevance: Insulin resistance plays a major role in causing type 2 diabetes mellitus (T2DM). This grant will examine the cellular mechanisms of insulin resistance in young lean insulin resistant offspring of parents with T2DM, who have a strong likelihood of developing T2DM using state-of-the-art magnetic resonance spectroscopy methods to non invasively examine intracellular metabolism. It is anticipated that the results of these studies will yield important new insights into the causes of insulin resistance, which will lead to new therapies for T2DM.